A large number of vectors for producing recombinant proteins have been developed, and the expression levels of proteins are high in expression systems that use bacteria such as E. coli, eukaryotic microorganisms such as yeast, and insect cells as host. However, when expressing proteins unique to mammals, they may not form a normal three-dimensional structure, and most of the time there is a problem with post-translational modifications such as glycosylation. Thus, it is necessary to establish expression systems that use mammalian cells as host, but in general, the expression level is low in most cases. Furthermore, expression systems that use recombinant virus vectors are also used in animal cells, which are higher than insect cells, but removing recombinant virus vectors from the expressed proteins is a very cumbersome process and the risk of virus vectors themselves cannot be denied.
Cases of recombinant protein production using a mammalian cell as host include tissue plasminogen activator (Patent Document 1), erythropoietin (Patent Document 2 and Non-patent Documents 1-3), IFN-γ (Non-patent Document 4), and IFN-β (Patent Document 3 and Non-patent Document 5). Furthermore, there are many reports about recombinant production of monoclonal antibodies (Patent Documents 4 to 6, and Non-patent Documents 6 to 8). In addition, an example of a high expression vector for mammalian cells is pNOW/CMV-AA (Patent Document 7). The production level of conglutinin using this vector was up to 11.8 μg/mL after four days of culture. However, the production level of recombinant protein is unlikely to be sufficient in these cases.
Prior art documents relating to the invention of this application are shown below.